Plasma Small Extracellular Vesicle-Carried miRNA-501-5p Promotes Vascular Smooth Muscle Cell Phenotypic Modulation-Mediated In-Stent Restenosis

Abstract

Vascular smooth muscle cell (VSMC) phenotypic modulation plays an important role in the occurrence and development of in-stent restenosis (ISR), the underlying mechanism of which remains a key issue needing to be urgently addressed. This study is designed to investigate the role of plasma small extracellular vesicles (sEV) in VSMC phenotypic modulation. sEV were isolated from the plasma of patients with ISR (ISR-sEV) or not (Ctl-sEV) 1 year after coronary stent implantation using differential ultracentrifugation. Plasma sEV in ISR patients are elevated markedly and decrease the expression of VSMC contractile markers α-SMA and calponin and increase VSMC proliferation. miRNA sequencing and qRT-PCR validation identified that miRNA-501-5p was the highest expressed miRNA in the plasma ISR-sEV compared with Ctl-sEV. Then, we found that sEV-carried miRNA-501-5p level was significantly higher in ISR patients, and the level of plasma sEV-carried miRNA-501-5p linearly correlated with the degree of restenosis ( $R^2=0.62$ ). Moreover, miRNA-501-5p inhibition significantly increased the expression of VSMC contractile markers α-SMA and calponin and suppressed VSMC proliferation and migration; in vivo inhibition of miRNA-501-5p could also blunt carotid artery balloon injury induced VSMC phenotypic modulation in rats. Mechanically, miRNA-501-5p promoted plasma sEV-induced VSMC proliferation by targeting Smad3. Notably, endothelial cells might be the major origins of miRNA-501-5p. Collectively, these findings showed that plasma sEV-carried miRNA-501-5p promotes VSMC phenotypic modulation-mediated ISR through targeting Smad3.